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With the aim of testing the hypothesis that increased ovulation number increases the frequency or number of ovarian cysts, the development and appearance of ovarian cysts was investigated in mice of differing ages and total lifetime ovulation number. High total ovulation number was induced by keeping mice in cages divided by a screen, with a male on one side and two females on the other side. Significantly more cysts were observed in animals subjected to incessant ovulation for 8 months and in 12 month breeding mice than in 3\u2010month virgin mice or 1\u2010month prepubertal animals. These cysts had the appearance of benign serous inclusion cysts. When cystic ovaries were serial sectioned, 47% of cysts had a connection to the ovarian hilus and potentially to the tubules of the rete ovarii, 31% were adjacent to the hilus, and 22% had an intra\u2010ovarian location. A significant increase in intra\u2010ovarian cysts was observed in the 8\u2010month incessant ovulation group, implying that high ovulation number leads to ovarian surface invagination and inclusion cyst formation. In conclusion, ovarian inclusion cysts may be derived from more than one epithelial source, but incessant ovulation may increase the proportion derived from the ovarian surface epithelium. Because the cysts observed resembled human serous inclusion cysts these results have possible implications for epithelial ovarian carcinoma. Copyright \u00a9 2005 Pathological Society of Great Britain and Ireland. 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